Oxidation of arachidonic acid (AA) generates a variety of biologically active mediators. Three distinct arachidonic acid metabolic pathways have been described. Of the three pathways, considerable attention has been paid to the products of the cyclooxygenase and lipoxygenase pathways; and therapeutic effects have been well described.
By contrast, much less attention has been paid to the products of the so-called “third pathway.” Unlike the other two pathways, this pathway is mediated by cytochrome P450 monooxygenases and uses NADPH and molecular oxygen in a 1:1 stoichiometry.
The metabolism of arachidonic acid by cytochrome P450 monoxygenases leads to the formation of various biologically active eicosanoids. Three types of oxidative reactions are known to occur. First, olefin epoxidation (catalyzed by epoxygenases) gives rise to the epoxyeicosatrienoic acids (EETs). Four important EET regioisomers are [5,6]-EET, [8,9]-EET, [11,12]-EET, and [14,15]-EET. The EETs are hydrolyzed by epoxide hydrolases to form the corresponding dihydroxycicosatrienoic acids (DHETs). Second, omega terminal oxidation leads to the formation of omega terminal hydroxyeicosatetraenoic acids (HETEs). Third, allylic oxidation leads to the formation of midchain HETEs.
Several cytochrome P450 epoxygenases have been identified, including members of the CYP1A, CYP2B, CYP2C, CYP2E, and CYP2J subfamilies. Attention has recently been focused on proteins of the CYP2J subfamily. One particular isoform, CYP2J2, is highly expressed in human cardiac myocytes, where arachidonic acid is metabolized to s produce EETs (Wu et al., 271 J. Biol. Chem. 12551 (1996)). CYP2J2 proteins are also found in epithelial cells in the airway and in the gut (Zeldin et al., 51 Mol. Pharm. 931 (1997); Zeldin et al., 50 Mol. Pharm. 1111 (1996)). In contrast to the other P450 enzymes, CYP2J2 proteins are distributed uniformly along the length of the gut, in epithelial and non-epithelial cells. High levels of the CYP2J2 proteins are found in cells of the autonomic ganglia, epithelial cells, and intestinal smooth muscle cells. Several CYP2J homologues have been identified in animals including rat CYP2J3, rat CYP2J4, mouse CYP2J5 and mouse CYP2J6 (Zhang et al., 340 Arch. Biochem. Biophys. 270 (1997); Ma et al., 274 J. Biol. Chem. 17777 (1999)).
The EETs are considered to be potential candidates for endothelium-derived hyperpolarizing factor (EDHF) because they hyperpolarize and relax vascular smooth muscle cells by activating calcium-sensitive potassium (KCa) channels (Campbell et al., 78 Circ. Res. 415 (1996); Rosolowsky et al., 1299 Biochim. Biophys. Acta. 267 (1996). EDHF is the substance that produces the vascular smooth muscle hyperpolarization which cannot be explained by nitric oxide (NO; the so-called endothelium-derived relaxing factor; EDRF). In the coronary microcirculation, EDHF and not NO is the predominant mediator of endothelium-dependent relaxation. EETs increase coronary blood flow and protect the myocardium from ischemia-reperfusion injury (Wu et al., 272 J. Biol. Chem 12551 (1997); Oltman et al., 83 Circ. Res. 932 (1998)).
What is not known, however, is whether EETs have an additional therapeutic usefulness.